A novel process for making an isoflavone concentrate product from soybeans which includes diluting solubles from alcohol-extracted hexane-defatted soybean flakes to about 10% to about 30% solids, separating undissolved solids from the diluted soy solubles, such that the separated solids have at least 4% isoflavones by weight of dry matter. That concentrate can then be further concentrated to at least 40% isoflavones by weight of dry matter by adjusting pH and temperature and extracting with solvents. The soy isoflavone concentrate products are then used in liquid or dry beverage, food or nutritional products.
This invention relates to a process for making an isoflavone concentrate product from soybeans. Isoflavones are a unique class of phytoestrogensxe2x80x94plant hormonesxe2x80x94that naturally occur in soybeans.
It is anticipated that consumer demand for soy isoflavones will continue to grow. Scientists have demonstrated that isoflavones have the ability to inhibit cancer cell growth, and some researchers believe that isoflavones may contribute to soy""s ability to lower blood-cholesterol levels.
Research shows that soy isoflavones have a wide range of health benefits that include moderating normal symptoms associated with menopause and promoting bone and heart health. It appears that about 100 milligrams of isoflavones (expressed in the glucoside form) are necessary to deliver most of these health benefits. This is about the average amount consumed daily by Asian men and women who have a much lower incidence of heart disease, osteoporosis and uncomfortable menopausal symptoms compared to Western societies.
Some women""s health problems during and after middle age are related to a changing hormonal state. Consuming soy isoflavones can help moderate the natural hormonal changes associated with several menopausal and postmenopausal symptoms.
Soy isoflavones are potent antioxidants capable of reducing the amount of LDL-cholesterol (bad cholesterol) that undergoes modification in the body. Entry of the modified LDL-cholesterol into the walls of blood vessels contributes to the formation of plaques. These plaques cause the blood vessels to lose their ability to function normally. Research in both animals and humans shows that ingesting soy isoflavones can help maintain normal blood vessel function.
Soy isoflavones are actively studied for their effects on maintaining and improving bone health. Women can lose up to 15% of their total bone mass in the early years following the onset of menopause. This loss can be quite detrimental, particularly to women who enter menopause with weaker bones. Emerging research shows that isoflavones appear to play a role in both preventing bone loss and increasing bone density.
The principal types of isoflavones found in soybeans are glucones (with sugars) and aglucones (without sugars). Glucones have the glucose molecule attached, and include genistin, daidzin and glycitin. Aglucones are isoflavones without the glucose molecule, and they include genistein, daidzein and glycitein. It is an object of this invention to produce products with the isoflavones genistein, daidzein and glycitein in similar proportions as those found naturally in soybeans when isoflavones are reported in the aglucone form.
The prior art teaches isolating genistin from hexane-extracted soybean flakes. Walter (xe2x80x9cGenistin (an Isoflavone Glucoside) and Its Aglucone, Genistein, from Soybeans,xe2x80x9d J. of Am. Chem. Soc., 63, 3273 (1941)) describes a method involving, among other steps, extracting the flakes with methanol, precipitating with acetone and recrystallizing with ethanol.
U.S. Pat. No. 5,141,746 (Fleury et al.) describes a method for preparing an impure extract of two specific isoflavones daidzin malonate and genistin malonate. Fleury describes a method involving, among other steps, mixing hexane-defatted ground soybeans with 80 percent (%) aqueous methanol, filtering and drying; adjusting pH multiple times with, among other chemicals, hydrochloric acid and sodium hydroxide, and extracting with an organic solvent, such as butanol.
U.S. Pat. No. 5,352,384 (Shen) describes making an aglucone enriched fiber. Shen describes solubilizing isoflavones from soy flour by, among other steps, forming a slurry with an extractant, such as sodium, potassium or calcium hydroxide, to adjust the pH to the proteins"" isoelectric point of 6.7-9.7, and reacting the slurry with the enzyme xcex2-glucosidase.
It is apparent that an efficient process for removing isoflavones from soybeans is needed. It also is apparent that a low-cost soy isoflavone concentrate (SIC) product is needed.
The invention uses the by-product soy solubles to SIC products. Soy solubles are recovered from alcohol-extracted hexane-defatted soybean flakes. These solubles, sometimes called soy xe2x80x9cmolassesxe2x80x9d, are desolventized, such that they contain less than 0.5% alcohol, and typically are evaporated to 60% solids.
It was discovered that soy solubles, on average, contain 3.31 milligrams per gram (mg/g) genistin on a wet basis and have a total isoflavone content (i.e., daidzin, glycitin, genistin, mal-daidzin, mal-genistin, daidzein, glycitein, genistein and some unidentified isoflavones) of 8.96 mg/g on a wet basis as determined by high performance liquid chromatography (HPLC). The genistin to daidzin ratio of soybeans is about 1 to 1 and that ratio in soy solubles was found to be about 1-1.5 to 1.
It was further discovered that if the soy solubles are diluted with water to form a slurry and the undissolved solids are removed from the slurry to form a wet xe2x80x9ccakexe2x80x9d, the cake contains a significantly concentrated amount of isoflavones.
It was further discovered that if the soy solubles, which have about 20 mg/g isoflavones on a dry basis, are diluted with water to a certain percent solids and the undissolved solids are separated from the diluted solubles that the isoflavone content of the solubles can be concentrated by at least 2 times, or about 4% isoflavones by weight of dry matter.
It was further discovered that the 4% isoflavone material can then be further concentrated to at least 40% isoflavones by weight of dry matter by adjusting pH and temperature and extracting with solvents, like acetone and hexane.
It was further discovered that acetone extraction of the isoflavone material separated from the soy solubles also yields a by-product solution that is high in saponins, which are beginning to be recognized as having many useful nutritional and health benefits like soy isoflavones.
The invention comprises a novel process for manufacturing a novel soy isoflavone concentrate (SIC) product. The isoflavone content by weight of the SIC product is 8 to 11 times that of soy flour, which typically has about 0.6% by weight isoflavones. More particularly, the SIC product contains at least 4% by weight isoflavones, 20-60% protein, with it typically being at least one-third protein, and a relatively low amount of fiber, with it typically being less than 5% crude fiber and less than 20% dietary fiber. The 4% material can then be further concentrated to produce a SIC product having at least 40% by weight isoflavones, with such product typically being greater than 80% carbohydrates, less than about 5-10% protein, about one-third dietary fiber and less than 5-10% crude fiber. The SIC products are then used in liquid or dry beverage, food or nutritional products.
The invention comprises a novel process for manufacturing a novel soy isoflavone concentrate (SIC) product. The isoflavone content by weight of the SIC product is 8 to 11 times that of soy flour, which typically has about 0.6% by weight isoflavones. More particularly, the SIC product contains at least 4% by weight isoflavones, 20-60% protein, with it typically being at least one-third protein, and a relatively low amount of fiber, with it typically being less than 5% crude fiber and less than 20% dietary fiber. The 4% material can then be further concentrated to produce a SIC product having at least 40% by weight isoflavones, with such product typically being greater than 80% carbohydrates, less than about 5-10% protein, about one-third dietary fiber and less than 5-10% crude fiber. The SIC products are then used in a liquid or dry beverage, food or nutritional product.
The steps of the subject invention for making the above-described 4% isoflavone concentrate material are: dehulling whole soybeans; flaking the dehulled soybeans; extracting soybean oil from the flaked soybeans with hexane, a solvent; desolventizing the defatted soybean flakes without high heating or toasting to produce xe2x80x9cwhitexe2x80x9d flakes; extracting the white flakes with aqueous alcohol; recovering solubles from the extraction; desolventizing (removing alcohol) from the soy solubles; diluting the soy solubles with water to form a slurry; separating the undissolved solids from the slurry to form a cake and drying the wet cake.
The steps of the subject invention for making the above-described 40% isoflavone concentrate material are: providing solubles from desolventized alcohol-extracted hexane-defatted soybean flakes; diluting said solubles with water to about 10-30% solids; separating undissolved solids from said diluted solubles; mixing said separated solids with acetone and sodium hydroxide, with said mixture having a pH of about 6.5-7; heating said mixture to about 55-60 degrees Celsius and holding said heated mixture for an effective period of time; separating a centrate from said mixture; cooling said centrate; decanting said acetone from said cooled centrate to form an acetone extract of said isoflavones; removing said acetone from said extract; raising the pH of said extract to about 10-10.5 with sodium hydroxide; heating said extract to about 50-60 degrees Celsius; lowering the pH of said extract to about 4.5-5 with hydrochloric acid; mixing said extract with hexane; heating said second mixture to about 50-60 degrees Celsius and stirring for an effective period of time; removing said hexane from said second mixture to form a second extract; separating solids from said second extract with chilling and drying said solids to form the product having about at least 40% isoflavones by weight.
The general procedure for extracting soybean oil from soybeans is well described in the prior art. E.g., xe2x80x9cExtraction of Oil from Soybeans,xe2x80x9d J. Am. Oil Chem. Soc., 58, 157 (1981) and xe2x80x9cSolvent Extraction of Soybeans,xe2x80x9d J. Am. Oil Chem. Soc., 55, 754 (1978).
An initial step in soybean oil extraction is the dehulling process in which the soybean hulls are removed from the whole soybeans. The soybeans are carefully cleaned prior to dehulling to remove foreign matter, so that product will not be contaminated by color bodies. Soybeans also are normally cracked into about 6 to 8 pieces prior to dehulling.
The hull typically accounts for about 8% of the weight of the whole soybean. The dehulled soybean is about 10% water, 40% protein, 20% fat, with the remainder mainly being carbohydrates, fiber and minerals.
The next step in soybean oil extraction involves the flaking process. Soybeans are conditioned prior to flaking by adjusting moisture and temperature to make the bean pieces sufficiently plastic. The conditioned bean pieces are passed through flaking rolls to form flakes about 0.01 to 0.012 inches (in.) thick.
The soybean flakes are defatted by contacting them with hexane to remove the soybean oil. Soybean oil is used in margarine, shortening and other food and products, and is a good source of lecithin, which has many useful applications as an emulsifier.
A detailed description of the general procedure for the alcohol process for manufacturing soy protein concentrate (SPC) is found in U.S. Pat. Nos. 3,365,440 (Circle et al.) and 5,097,017 (Konwinski). SPC has been described in commerce as a product containing not less than 70% protein (Nxc3x976.25). See A. K. Smith and S. J. Circle, Editors, xe2x80x9cSoybeans: Chemistry and Technology, Volume I, Proteins,xe2x80x9d the AVI Publishing Co., 1973.
In the alcohol process, the hexane-defatted soybean flakes are desolventizedxe2x80x94hexane is removedxe2x80x94without toasting to produce white flakes. This is different than conventional soybean oil hexane processes where the flakes are toasted and used for animal feed. Instead of being further processed into SPC, the white flakes can be ground to make soy flour.
The white flakes are extracted with 55-75%, typically 60%, by weight aqueous ethanol in a countercurrent (flake to solvent flow) extraction devicexe2x80x94extractor. The alcohol to flake ratio is about 5 to 1.
The alcohol extraction removes carbohydrates, including oligosaccharides, from the white flakes, which thereby increases the protein content of the material. A typical sample of soy molasses from the SPC alcohol process was found to contain 7.80, 128.50, 19.45 and 86.79 milligrams/gram (mg/g) glucose, sucrose, raffinose and stachyose, respectively, on a wet basis. Soy molasses also typically contains 7-8% protein.
In the preferred embodiment of this invention, the diluting, separating, pasteurizing and drying steps to make the 4% isoflavone product are performed in a continuous process.
The soy solubles are diluted with water to form a slurry. Cold tap water is preferred source of water for the dilution.
In a preferred embodiment of this invention, the solubles are diluted to about 10% to about 30%, most preferred 18% solids. The slurry""s pH being 5.5-6.
It also usually is necessary to provide some agitation or mixing to slurry the diluted solubles. One means for performing the mixing is a propeller-type agitator. The diluted soy solubles may be optionally cooled using a chiller.
The undissolved solids are removed from the slurry to form a wet cake. The undissolved solids could be removed by a number of physical separation means; however, centrifugation is the most efficient and effective means.
In the preferred embodiment of this invention, a scroll-type centrifuge is used to remove the undissolved solids from the slurry, such that wet cake is 25-30% solids and contains about 10-20% of the soy solubles"" solids. In yet another embodiment of this invention, the separation can be performed with a disc type or tubular centrifuge.
Alternatively, the dilution and separation steps 8 and 9 could be described as water xe2x80x9cwashingxe2x80x9d the soy solubles. These steps serve to concentrate the soy isoflavones from the solubles. In another embodiment of this invention, the washing process may be repeated one or more times in an effort to further concentrate the isoflavones, however, product yield (the quantity of SIC produced) would decrease.
In a preferred embodiment of this invention the wet cake is pasteurized prior to the drying step, so that the SIC will test negative for salmonella and have an acceptable microbial profile. One means for pasteurization is to hold the wet cake in a steam-jacketed kettle for 10 minutes at 160 degrees Fahrenheit (xc2x0F.).
The wet cake is dried to produce SIC useable as a nutritional supplement, or a food ingredient or product. The preferred means of drying is a vertical spray drier with a high pressure nozzle. To facilitate spray drying, the wet cake is diluted to less than 25% solids, most preferred 15%, prior to pasteurization.
A lot prepared by the above method had 6.9% isoflavones (66.7% genistein; 32.1% daidzein and 1.2% glycitein when isoflavones are reported in the aglucone form); 32.8% protein; 2.1% fat; 55.6% total carbohydrates; 4.4% moisture and 5.1% ash, with no heavy metals detected. When analyzed for isoflavones using standard HPLC methodology, the lot had 40.7 milligrams/gram (mg/g) genistin; 20.7 mg/g daidzin; 0.8 mg/g glycitin; 1 mg/g daidzein; 0.7 mg/g genistein and 0 glycitein.
Another lot prepared by the above method had 4.5% isoflavones (57.8% genistein; 37.4% daidzein and 4.6% glycitein when isoflavones are reported in the aglucone form); 30.2% protein; 1.3% fat; 59.5% total carbohydrates; 3.8% moisture and 5.2% ash, with no heavy metals detected. When analyzed for isoflavones using standard HPLC methodology, the lot had 23.7 milligrams/gram (mg/g) genistin; 14.3 mg/g daidzin; 2.0 mg/g glycitin; 1.6 mg/g daidzein; 0.04 mg/g genistein and 0 glycitein. These about 4% isoflavone concentrate products typically contain about 10% saponins by weight.
The spray-dried powdered SIC has many uses. For example, it can be tableted or used in drink mixes.
The spray-dried powdered SIC may be coated with commercial lecithin or other food-grade surfactants, such as mono-diglycerides, to improve water dispersibility and reduce clumping of the product. Such a coating-addition would be in the range of about 0.5%.
Prior to or after spray drying the wet cake, the cake can be further concentrated to at least 40% by weight isoflavones by adjusting pH and temperature and extracting with solvents.
The wet cake or slurried spray-dried cake is mixed with a solvent and its pH is adjusted to about 6-7, most preferred 6.4-6.8. Soy solubles or diluted soy solubles could also be suitable materials for this isoflavone extraction process.
Acetone is the preferred solvent. 50% sodium hydroxide is the preferred base for adjusting pH if necessary.
The ratio of starting material to the acetone/water mixture can vary. The efficiency of extraction using a ratio of starting material to acetone/water mixture of approximately 1 to 10 by mass is typically very high, often exceeding 90%.
The mixture is then extracted by heating to acetone reflux, 57-58 degrees Celsius. Alternatively, the extraction could be performed without heating if given sufficient time.
In addition to heating, it also preferred to hold the material for about 90 minutes to aid the extraction. It also preferred to then cool the heated material.
After extraction, a centrate is separated from the mixture. One means of separation is with a horizontal type centrifuge. Solids from the centrifugation may be dried with a pin dryer. The solids contain the majority of the protein in the about 4% isoflavone starting material.
After separation, it is desired to cool the centrate. One means for cooling the centrate is by passing it through a heat exchanger.
Saponins, another phytochemical, can then be removed from the centrate by decanting the solvent. The majority, at least about 60%, of the saponins in the 4% isoflavone starting material are in the solution. The isoflavones remain in the decanted solvent. It is preferred to cool and hold the centrate at less than 10 degrees Celsius prior to decanting to accumulate the saponins.
The saponins can be separated from the solution. One means of separation is with a disc type centrifuge.
The solvent, acetone in the preferred embodiment, is then removed from the decanted solvent. One means for removing the solvent is stripping it at ambient temperature to 90-95 degrees Celsius to form an extract. The recovered alcohol can then be re-used in the process.
The next steps in the process serve to saponify and remove lipids and certain emulsifying agents, like lecithin. This facilitates drying of the isoflavone concentrate.
The pH of the extract is then raised, most preferred 10-10.5. Again, sodium hydroxide is a suitable reagent for raising pH. It is then preferred to heat the pH raised extract to 60 degrees Celsius. These steps cleave glycerides from the lipids.
The pH of the extract is then further adjusted by lowering the pH, most preferred 4.5-5. Hydrochloric acid is a suitable reagent for lowering the pH. The chlorine binds with sodium in the extract to form sodium chloride.
An alkane is then used to perform a second extraction. Hexane is the preferred alkane. If necessary, water is added prior to adding the alkane. Free fatty acids are extracted by the hexane.
The alkane mixture is then heated to aid extraction, most preferred about 50 degrees Celsius. It also preferred to vigorously stir the mixture for about 30 minutes.
The mixture is then decanted by allowing it to settle. The decanted hexane and rag layers can then be distilled, so that the hexane may be re-used in the process.
The decanted water layer could then be dried. It is, however, preferred to chill the decanted water layer with precipitate to 5 degrees Celsius.
Wet solids are then separated with a disc type centrifuge. Secondary recovery of wet solids from the centrate of that centrifugation can be done with a horizontal type centrifuge to increase yield. The centrate from the centrifugation contains soluble carbohydrates from the starting material.
The wet solids are typically dried with a contact freeze drier. Prior to drying, the wet solids are sometimes blended or diluted with other isoflavone materials, such soy solubles or the above-described 4% material, to achieve a desired isoflavone content.
A lot prepared by the above method had 55.6% isoflavones (62.5% genistein; 34% daidzein and 3.5% glycitein when isoflavones are reported in the aglucone form); 5.4% protein; 1.1% fat; 87.2 total carbohydrates; 35.1% dietary fiber; 8.9% total sugars; 2.0% moisture and 4.3% ash. When analyzed for isoflavones using standard HPLC methodology, the lot had 278.9 milligrams/gram (mg/g) genistin; 150.4 mg/g daidzin; 15.9 mg/g glycitin; 2.9 mg/g daidzein; 2.6 mg/g genistein and 0 glycitein.
Another lot prepared by the above method had 49.3% isoflavones (72.7% genistein; 25.3% daidzein and 2.4% glycitein when isoflavones are reported in the aglucone form); 1.8% protein; 2.0% fat; 89.6% total carbohydrates; 2.4% moisture and 5.1% ash. When analyzed for isoflavones using standard HPLC methodology, the lot had 333.5 milligrams/gram (mg/g) genistin; 117.2 mg/g daidzin; 11.3 mg/g glycitin; 3.2 mg/g daidzein; 3.4 mg/g genistein and 0 glycitein. These isoflavone concentrates typically contain about 10-20% saponins by weight.
These and other aspects of the present invention may be more readily understood by reference to one or more of the following examples. These examples illustrate the practice of this invention.